title: Optimization of experimental design parameters for high-throughput chromatin immunoprecipitation studies.
abstract: High-throughput, microarray-based chromatin immunoprecipitation (ChIP-chip) technology allows in vivo elucidation of transcriptional networks. However this complex is not yet readily accessible, in part because its many parameters have not been systematically evaluated and optimized. We address this gap by systematically assessing experimental-design parameters including antibody purity, dye-bias, array-batch, inter-day hybridization bias, amplification method and choice of hybridization control. The combined performance of these optimized parameters shows a 90% validation rate in ChIP-chip analysis of Myc genomic binding in HL60 cells using two different microarray platforms. Increased sensitivity and decreased noise in ChIP-chip assays will enable wider use of this methodology to accurately and affordably elucidate transcriptional networks.
authors.affiliation: Division of Cancer Genomics and Proteomics, Ontario Cancer Institute, University Health Network, University of Toronto, Toronto, ON, Canada.
authors.forename: Romina
authors.initials: R
authors.lastname: Ponzielli
authors.forename: Paul C
authors.initials: PC
authors.lastname: Boutros
authors.forename: Sigal
authors.initials: S
authors.lastname: Katz
authors.forename: Angelina
authors.initials: A
authors.lastname: Stojanova
authors.forename: Adam P
authors.initials: AP
authors.lastname: Hanley
authors.forename: Fereshteh
authors.initials: F
authors.lastname: Khosravi
authors.forename: Christina
authors.initials: C
authors.lastname: Bros
authors.forename: Igor
authors.initials: I
authors.lastname: Jurisica
authors.forename: Linda Z
authors.initials: LZ
authors.lastname: Penn
mesh_terms: D000906:Antibodies / Q000276:immunology; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D018922:HL-60 Cells; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D016271:Proto-Oncogene Proteins c-myc / Q000276:immunology / Q000378:metabolism; D050436:Regulatory Elements, Transcriptional
chemical_list: D000906:Antibodies; D016271:Proto-Oncogene Proteins c-myc